Over 300,000 infants are born with sickle cell disease (SCD) every year world-wide, including at least 1,000 in the US. Prenatal diagnosis by amniocentesis or chorionic villus sampling is available; but high cost, invasiveness, and risk of miscarriage limit their use. This Phase I SBIR project will show feasibility of a non- invasive test for Sickle Cell Disease that is affordable and completely safe to use. By finding their child?s SCD status before birth, parents can opt for umbilical cord blood banking to save their child?s stem cells. These stem cells could be critical for access to future gene-therapy SCD cures that are in clinical trials and will likely become approved in the next 3-5 years. Our non-invasive prenatal test uses the cell-free DNA that a developing fetus sheds into the mother?s bloodstream. By taking peripheral blood sample from a pregnant woman, our assay is able to determine the fetal genotype for sickle cell disease causing mutations. To support this assay, we have developed innovative probes for sickle cell alleles that can measure allele fractions with coefficient of variation <2% through the use of next- generation DNA sequencing. This project will demonstrate the feasibility of our approach for non-invasive prenatal testing of sickle cell disease through the following specific aims. Specific Aim 1: Validate optimized probes for HBB on cfDNA isolated from healthy SCT individuals. Specific Aim 2: Identify the fetal HBB genotype (e.g. HbAS, HbSC, or HbSS) from pregnant women who have SCD. Aim 3: Identify the fetal SCD genotype (e.g. HbAA, HbAS, HbSC, or HbSS) of pregnant women who have SCT.